The present invention relates generally to the fields of biochemistry and medicine. In particular, the present invention relates to methods for use in delivering one or more exogenous polynucleotide sequences to muscle cells of an animal, in particular a mammal, so as to achieve an enhanced therapeutic effect thereon.
Various approaches have been suggested for the introduction of exogenous polynucleotide sequences into human or animal patients (generally referred to as "gene therapy"). Various techniques for integration of exogenous DNA into the genome of a cell have been proposed. Unfortunately, such integration can cause damage to the genome and possible genetic changes in the recipient cell which might predispose the cells to malignancy. Therefore, methods which avoid these potential problems would be desirable in providing safe and effective techniques of gene therapy.
The introduction of plasmid DNA into striated muscles by direct injection has been described in limb, trunk, and heart [Wolff, J. A. et at. (1990) Science 247:1465-1468; Lin, H. et al. (1990) Circulation 82:2217-2221; Acid, G. et at. (1991) New Biol. 3:71-81; Kitsis, R. et al. (1991) Proc. Natl. Acad. Sci. U.S.A. 88:4138-4142]. In addition, PCT published application WO 90/11092 (the entire disclosure of which is hereby incorporated by reference) generally describes the delivery of a purified polynucleotide to the interior of a cell or cells of a vertebrate, wherein the delivered material consists of a pharmaceutically acceptable carrier and a naked polynucleotide which encodes either a polypeptide or an antisense polynucleotide. Although a variety of types of cells (including, in particular, muscle cells) are suggested as target cells, there is no specific disclosure of the injection of polynucleotide into the tongue. WO 90/11092 proposes that the polynucleotide is integrated into the genomic DNA of the cell or cells and subsequently produces a messenger RNA which serves to encode a polypeptide with either pharmaceutical or immunogenic properties. The polypeptide may be released into the interstitial spaces surrounding the cell or function inside the cell. Alternatively, the integrated polynucleotide encodes a RNA sequence which inhibits in a cell the production of a polypeptide which is associated with a detrimental effect on either the cell or the entire organism. The methods described in WO 90/11092 are alleged to result in either immunogenic or pharmacological effects on the vertebrate through several means, including delivering a therapeutic polypeptide to the cells of the vertebrate and providing a transitory method of gene therapy. Although the direct injection of DNA is a relatively simple methodology, the levels of expression achieved to date have not at all been satisfactory for reliable achievement of therapeutic or other physiological effects on mammals so treated.
It is an object of the present invention to provide methods and compositions for achieving enhanced levels of expression of exogenous polynucleotide in animals upon direct injection of the polynucleotide into the mammal.